Dynamic regulation of the mitochondrial network by mitofusins (MFNs) modulates energy production, cell survival, and many intracellular signaling events, including calcium handling. However, the relative importance of specific mitochondrial functions and their dependence on MFNs vary greatly among cell types. Osteoclasts have many mitochondria, and increased mitochondrial biogenesis and oxidative phosphorylation enhance bone resorption, but little is known about the mitochondrial network or MFNs in osteoclasts. Because expression of each MFN isoform increases with osteoclastogenesis, we conditionally deleted MFN1 and MFN2 (double conditional KO (dcKO)) in murine osteoclast precursors, finding that this increased bone mass in young female mice and abolished osteoclast precursor differentiation into mature osteoclasts in vitro. Defective osteoclastogenesis was reversed by overexpression of MFN2 but not MFN1; therefore, we generated mice lacking only MFN2 in osteoclasts. MFN2-deficient female mice had increased bone mass at 1 year and resistance to Receptor Activator of NF-κB Ligand (RANKL)-induced osteolysis at 8 weeks. To explore whether MFN-mediated tethering or mitophagy is important for osteoclastogenesis, we overexpressed MFN2 variants defective in either function in dcKO precursors and found that, although mitophagy was dispensable for differentiation, tethering was required. Because the master osteoclastogenic transcriptional regulator nuclear factor of activated T cells 1 (NFATc1) is calcium-regulated, we assessed calcium release from the endoplasmic reticulum and store-operated calcium entry and found that the latter was blunted in dcKO cells. Restored osteoclast differentiation by expression of intact MFN2 or the mitophagy-defective variant was associated with normalization of store-operated calcium entry and NFATc1 levels, indicating that MFN2 controls mitochondrion–endoplasmic reticulum tethering in osteoclasts.

Studies from global loss‐of‐function mutants suggest that alternative NF‐κB downstream of NF‐κB inducing kinase (NIK) is a cell‐intrinsic negative regulator of osteogenesis. However, the interpretation of the osteoblast and/or osteocyte contribution to the bone phenotype is complicated by simultaneous osteoclast defects in these models. Therefore, we turned to a transgenic mouse model to investigate the direct role of NIK in the osteolineage. Osx‐Cre;NT3 animals (NT3‐Cre +), which bear a constitutively active NIK allele (NT3) driven by Osx‐Cre, were compared with their Cre‐negative, Control (Ctrl) littermates. NT3‐Cre + mice had elevated serum P1NP and CTX levels. Despite this high turnover state, µCT showed that constitutive activation of NIK resulted in a net increase in basal bone mass in both cortical and cancellous compartments. Furthermore, NT3‐Cre + mice exhibited a greater anabolic response following mechanical loading compared with controls. We next performed RNA‐Seq on nonloaded and loaded tibias to elucidate possible mechanisms underlying the increased bone anabolism seen in NT3‐Cre + mice. Hierarchical clustering revealed two main transcriptional programs: one loading‐responsive and the other NT3 transgene‐driven. Gene ontology (GO) analysis indicated a distinct upregulation of receptor, kinase, and growth factor activities including Wnts, as well as a calcium‐response signature in NT3‐Cre + limbs. The promoters of these GO‐term associated genes, including many known to be bone‐anabolic, were highly enriched for multiple κB recognition elements (κB‐RE) relative to the background frequency in the genome. The loading response in NT3‐Cre + mice substantially overlapped (>90%) with Ctrl. Surprisingly, control animals had 10‐fold more DEGs in response to loading. However, most top DEGs shared between genotypes had a high incidence of multiple κB‐RE in their promoters. Therefore, both transcriptional programs (loading‐responsive and NT3 transgene‐driven) are modulated by NF‐κB. Our studies uncover a previously unrecognized role for NF‐κB in the promotion of both basal and mechanically stimulated bone formation. © 2019 American Society for Bone and Mineral Research.

Conjunctival fibrosis remains the major impediment to the success of glaucoma filtration surgery. Anti-metabolites remain the gold standard for mitigating post-surgical fibrosis, but they are associated with high complication rates and surgical failure rates. Establishing a more targeted approach to attenuate conjunctival fibrosis may revolutionize the surgical approach to glaucoma. A new strategy is needed to prevent progressive tissue remodeling and formation of a fibrotic scar, subsequently increasing surgical success and reducing the prevalence of glaucoma-related vision loss. Advancements in our understanding of molecular signaling and biomechanical cues in the conjunctival tissue architecture are broadening the horizon for new therapies and biomaterials for the mitigation of fibrosis. This review aims to highlight the strategies and current state of promising future approaches for targeting fibrosis in glaucoma filtration surgery.

Bone infections, also known as infectious osteomyelitis, are accompanied by significant inflammation, osteolysis, and necrosis. Osteoclasts (OCs) are the bone-resorbing cells that work in concert with osteoblasts and osteocytes to properly maintain skeletal health and are well known to respond to inflammation by increasing their resorptive activity. OCs have typically been viewed merely as effectors of pathologic bone resorption, but recent evidence suggests they may play an active role in the progression of infections through direct effects on pathogens and via the immune system. This review discusses the host- and pathogen-derived factors involved in the in generation of OCs during infection, the crosstalk between OCs and immune cells, and the role of OC lineage cells in the growth and survival of pathogens, and highlights unanswered questions in the field.

Osteomyelitis can result from the direct inoculation of pathogens into bone during injury or surgery, or from spread via the bloodstream, a condition called hematogenous osteomyelitis (HOM). HOM disproportionally affects children, and more than half of cases are caused by Staphylococcus (S.) aureus . Laboratory models of osteomyelitis mostly utilize direct injection of bacteria into the bone or the implantation of foreign material, and therefore do not directly interrogate the pathogenesis of pediatric hematogenous osteomyelitis. In this study, we inoculated mice intravenously and characterized resultant musculoskeletal infections using two strains isolated from adults (USA300-LAC and NRS384) and five new methicillin-resistant S. aureus isolates from pediatric osteomyelitis patients. All strains were capable of creating stable infections over five weeks, although the incidence varied. Micro-computed tomography (microCT) analysis demonstrated decreases in trabecular bone volume fraction but little effect on bone cortices. Histologic assessment revealed differences in the precise focus of musculoskeletal infection, with varying mixtures of bone-centered osteomyelitis and joint-centered septic arthritis. Whole genome sequencing of three new isolates demonstrated distinct strains, two within the USA300 lineage and one USA100 isolate. Interestingly, this USA100 isolate showed a distinct predilection for septic arthritis, compared to the other isolates tested, including NRS384 and LAC, which more frequently led to osteomyelitis or mixed bone and joint infections. Collectively, these data outline the feasibility of using pediatric osteomyelitis clinical isolates to study the pathogenesis of HOM in murine models and lay the groundwork for future studies investigating strain-dependent differences in musculoskeletal infection.

Osteomyelitis can result from the direct inoculation of pathogens into bone during injury or surgery, or from spread via the bloodstream, a condition called hematogenous osteomyelitis (HOM). HOM disproportionally affects children, and more than half of cases are caused by Staphylococcus (S.) aureus. Laboratory models of osteomyelitis mostly utilize direct injection of bacteria into the bone or the implantation of foreign material, and therefore do not directly interrogate the pathogenesis of pediatric hematogenous osteomyelitis. In this study, we inoculated mice intravenously and characterized resultant musculoskeletal infections using two strains isolated from adults (USA300-LAC and NRS384) and five new methicillin-resistant S. aureus isolates from pediatric osteomyelitis patients. All strains were capable of creating stable infections over five weeks, although the incidence varied. Micro-computed tomography (microCT) analysis demonstrated decreases in trabecular bone volume fraction but little effect on bone cortices. Histologic assessment revealed differences in the precise focus of musculoskeletal infection, with varying mixtures of bone-centered osteomyelitis and joint-centered septic arthritis. Whole genome sequencing of three new isolates demonstrated distinct strains, two within the USA300 lineage and one USA100 isolate. Interestingly, the USA100 strain showed a distinct predilection for septic arthritis, compared to the USA300 strains, including NRS384 and LAC, which more frequently led to osteomyelitis or mixed bone and joint infections. Collectively, these data outline the feasibility of using pediatric osteomyelitis clinical isolates to study the pathogenesis of HOM in murine models and lay the groundwork for future studies investigating strain-dependent differences in musculoskeletal infection.